The present invention relates to molecular variants of the angiotensinogen gene. The present invention further relates to the diagnosis of these variants for the determination of a predisposition to hypertension, the determination of the prognosis of the predisposition to hypertension, and the management of hypertension.
The publications and other materials used herein to illuminate the background of the invention, or provide additional details respecting the practice, are incorporated by reference herein, and for convenience are respectively grouped in the appended List of References.
Hypertension is a leading cause of human cardiovascular morbidity and mortality, with a prevalence rate of 25-30% of the adult Caucasian population of the United States (JNC Report, (1985). The primary determinants of essential hypertension, which represents 95% of the hypertensive population, have not been elucidated in spite of numerous investigations undertaken to clarify the various mechanisms involved in the regulation of blood pressure. Studies of large populations of both twins and adoptive siblings, in providing concordant evidence for strong genetic components in the regulation of blood pressure (Ward (1990)), have suggested that molecular determinants contribute to the pathogenesis of hypertension.
Among a number of factors for regulating blood pressure, the renin-angiotensin system plays an important role in salt-water homeostasis and the maintenance of vascular tone; stimulation or inhibition of this system respectively raises or lowers blood pressure (Hall et al. (1990)), and may be involved in the etiology of hypertension. The renin-angiotensin system includes the enzymes renin and angiotensin-converting enzyme and the protein angiotensinogen (AGT). Angiotensinogen is the specific substrate of renin, an aspartyl protease. The structure of the AGT gene has been characterized (Gaillard et al. (1989); Fukamizu et al. (1990)).
Plasma angiotensinogen is primarily synthesized in the liver under the positive control of estrogens, glucocorticoids, thyroid hormones, and angiotensin II (Clauser et al.(1989)) and secreted through the constitutive pathway. Cleavage of the amino-terminal segment of angiotensinogen by resin releases a decapeptide prohormone, angiotensin-I, which is further processed to the active octapeptide angiotensin II by the dipeptidyl carboxypeptidase angiotensin-converting enzyme (ACE). Cleavage of angiotensinogen by renin is the rate-limiting step in the activation of the renin angiotensin system (Sealey et al. (1990)). Several observations point to a direct relationship between plasma angiotensinogen concentration and blood pressure; (1) a direct positive correlation (Walker et al. (1979)); (2) high concentrations of plasma angiotensinogen in hypertensive subjects and in the offspring of hypertensive parents compared to normotensives (Fasola et al. (1968)); (3) association of increased plasma angiotensinogen with higher blood pressure in offspring with contrasted parental predisposition to hypertension (Watt et al. (1992)); (4) decreased or increased blood pressure following administration of angiotensinogen antibodies (Gardes et al. (1982)) or injection of angiotensinogen (Menard et al. (1991)); (5) expression of the angiotensinogen gene in tissues directly involved in blood pressure regulation (Campbell and Habener (1986)); and (6) elevation of blood pressure in transgenic animals overexpressing angiotensinogen (Ohkubo et al. (1990; Kimura et al. (1992)).
The etiological heterogeneity and multifactorial determination which characterize diseases as common as hypertension expose the limitations of the classical genetic arsenal. Definition of phenotype, model of inheritance, optimal familial structures, and candidate-gene vs. general-linkage approaches impose critical strategic choices (Lander et al. (1986; White et al. (1987; Lander et al. (1989; Lalouel (1990; Lathrop et al. (1991)). Analysis by classical likelihood ratio methods in pedigrees is problematic due to the likely heterogeneity and the unknown mode of inheritance of hypertension. While such approaches have some power to detect linkage, their power to exclude linkage appears limited. Alternatively, linkage analysis in affected sib pairs is a robust method which can accommodate heterogeneity and incomplete penetrance, does not require any a priori formulation of the mode of inheritance of the trait and can be used to place upper limits on the potential magnitude of effects exerted on a trait by inheritance at a single locus. (Blackwelder et al. (1985; Suarez et al. (1984)).
Prior studies have it was found that the angiotensinogen gene is involved in the pathogenesis of essential hypertension. The following were found: (1) genetic linkage between essential hypertension and AGT in affected siblings; (2) association between hypertension and certain molecular variants of AGT as revealed by comparison between cases and controls; (3) increased concentrations of plasma angiotensinogen in hypertensive subjects who carry a common variant of AGT strongly associated with hypertension; (4) persons with the most common AGT gene variant exhibited not only raised levels of plasma angiotensinogen but also higher blood pressure; and (5) the most common AGT gene variant was found to be statistically increased in women presenting preeclampsia during pregnancy, a condition occurring in 5-10% of all pregnancies. The association between renin, ACE or AGT and essential hypertension was studied using the affected sib pair method (Bishop et al. (1990)) on populations from Salt Lake City, Utah and Paris, France, as described in further detail in the Examples. Only an association between the AGT gene and hypertension was found. The AGT gene was examined in persons with hypertension, and at least 15 variants have been identified. None of these variants occur in the region of the AGT protein cleaved by either renin or ACE. Identification of the AGT gene as being associated with essential hypertension was confirmed in a population study of healthy subjects and in women presenting preeclampsia during pregnancy. See, e.g., U.S. Pat. Nos. 5,374,525 and 5,763,168, each incorporated herein by reference; U.S. patent application Ser. No. 09/106,216, filed Jun. 29, 1998, incorporated herein by reference; Jeunemaitre et al. (1992); Jeunemaitre et al. (1993); and Jeunemaitre et al. (1997).
According to Gaillard et al. (1989), the human AGT gene contains five exons and four introns which span 13 Kb. The first exon (37 bp) codes for the 5' untranslated region of the mRNA. The second exon codes for the signal peptide and the first 252 amino acids of the mature protein. Exons 3 and 4 are shorter and code for 90 and 48 amino acids, respectively. Exon 5 contains a short coding sequence (62 amino acids) and the 3'-untranslated region. Genbank accession No. AH002594 also sets forth a sequence of the AGT gene as revised on Oct. 30, 1994. The revised sequence moves the start site of transcription one nucleotide 5' of the transcription start site identified in Gaillard et al. (1989). Since polymorphisms described herein and in the prior art have been written with respect to the Gaillard et al. (1989) transcription start site, this nomenclature will also be used herein.
It is an object of the present invention to identify additional AGT polymorphisms associated with hypertension and to utilize such polymorphisms for determining predisposition to hypertension in individuals. Identification of individuals who may be predisposed to hypertension will lead to better management of the disease.